The availability of a truely tissue specific growth promotor would provide a unique opportunity to investigate a wide variety of basic phenomena, including the control mechanisms involved in normal and malignant tissue growth and the role of growth promotors in the response to various toxins and tumor promotors. I have previously reported that normal weanling and regenerating adult rat livers contain a substance(s) (HSS) that stimulates liver growth in vivo and in vitro; an increase in the rate of DNA synthesis and an actual increase in cell number are observed and this stimulation is specific for the liver. The active material has been purified approximately 1700-fold by a combination of heat, ethanol and ammonium sulfate precipitation and ion exchange and gel filtration chromatography. The active material exhibits a MW of 20-25,000 and produces its effect only after a significant lag period, during which new RNA and protein must be synthesized in order for the stimulation of DNA synthesis to occur. The objectives of the current proposal are to complete purification of HSS to homogeneity and examine its mode of action. Purification will utilize isoelectric focusing, lectin columns, cyto-affinity and gel electrophoresis techniques. HTC cells will serve as the primary screening assay, due to their marked sensitivity to HSS. However, active fractions will be tested on normal hepatocytes in culture and in vivo. A purified material will allow development of an immune assay and identification of the tissue source of HSS, its presence in various growth states (normal and malignant) and its possible production in response to toxins and tumor promotors. Important questions concerning the time course of HSS stimulation, necessity for de novo protein and RNA synthesis, cell cycle specificity and effects on protein synthesis and degradation can be approached with the present, 1700-fold enriched preparation. HSS will also be radioiodinated to determine if the specificity of its action and the variability of cellular responses to HSS are due to differences in binding to a specific receptor for HSS.